1. Field of the Invention
The present invention relates to methods and devices for detecting biological entities and components associated with hypersensitivity reactions in patients with food allergies, autoimmunity or cancer.
2. Background
It is well established that food-related allergies cause a variety of illnesses, whether it be in humans or other animals. Approximately two percent of adults and about five percent of infants and young children in the United States suffer from food allergies and each year roughly 30,000 individuals require emergency room treatment as a result. At present, there is no cure for food allergies and a food allergic consumer must avoid the food to which the consumer is allergic. The timing and location of an allergic reaction to food is affected by digestion. For example, an allergic person may first experience a severe itching of the tongue or “tingling lips.” Vomiting, cramps or diarrhea may follow. Later, as allergens enter the bloodstream and travel throughout the body, they can cause a drop in blood pressure, hives or eczema, or asthma when they reach the lungs. The onset of these symptoms may vary from a few minutes to an hour or two after the food is eaten. Delayed reactions take hours or days to manifest symptoms.
Von Pirquet first described serum sickness, the prototype of Immune Complex disease in 1925. Any food protein entering the circulation in sufficient quantity can produce symptom patterns resembling serum sickness. If antigens make it into the blood stream, they can stimulate the production of antibodies. These antibodies can then combine with antigens in the blood stream to produce circulating immune complexes (CICs).
Food-enriched blood, coming from the gastrointestinal tract (GIT), goes through the liver where most immune-complexes are removed. If circulating complexes pass the liver filter, they may cause disturbances in many organs. The other path of absorption of molecules from the GIT is through lymphatic drainage. The lymph channels flow together to form the thoracic duct, a flimsy vessel which drains its contents into the subclavian vein. This pathway may direct antigenic molecules directly to the lungs where food antigens may excite intrinsic asthmatic attacks, bronchitis, or more serious and enigmatic inflammatory lung diseases.
The combination of antibody with antigen in the blood stream is a circulating immune complex (CIC). In most cases, CICs are simply removed from the circulation by macrophages prior to triggering a cascade of events which may cause multiple symptoms, and possibly tissue damage.
CIC's activate complement which is a circulating system of 25 proteins which interact to produce a variety of defensive molecular weapons. There are two main functions of the complement cascade. The first is to opsonize bacteria, viruses and antibodies with covalently bound C3b. The bacterial, viral or CIC-C3b complex binds to the CR1 receptors through the ligand C3b. CR1 receptors are found on red blood cells or other cells, such as macrophages which result in rapid removal of the C3b-CICs. The CR1 receptor is a cofactor that causes rapid degradation of C3b by Factor H and Factor I to CIC-C3bi and ultimately to CIC-C3d/C3d,g. It is noteworthy that C3d/C3d,g contain a thioester bond, which causes this fragment to remain covalently bound to the activator i.e. CIC, indefinitely. The second function is to lyse cells by activation of the terminal pathway proteins C5 through C9. C5-C9 attach to cell surfaces, assemble into pores (membrane attack complex), and disrupt the cell membrane or cell walls. The net effect is that ions and water flow into a cell causing the cell to burst.
Clearly the inadvertent or inappropriate activation of complement can have serious consequences for healthy self-cells and tissues. CIC's leave capillaries to trigger inflammatory events in target tissues. A classic model of complex-induced pathology is the Arthus reaction, which appears 3-6 hours after antigen challenge and involves large insoluble complexes with complement (C3b) passing through vessel walls to excite inflammatory responses in target tissues.
Regardless of the animal, allergens (antigens) from food, food additives or environmental sources cause an acquired immunity. Acquired immunity is simply the ability of allergens to either cause the production of antibodies (IgM, IgA, IgG, IgE and IgD) or interact with the mucosa or epidermis and stimulate T-cells. These antibodies react with the allergen and cause symptoms associated with allergy. Allergic reactions are classified into four types (I, II, III, IV) based on the Gell/Coombs scheme.
Immunoenzymometric assays involve the binding of an analyte of interest with a reaction or binding partner, where the binding partner carries a label. The binding partner is contained in a test strip, well or other apparatus so that it is non-reactive unless and until its partner analyte contacts the test strip. When this happens, the analyte and labelled binding partner bind to each other, forming a complex. Detection is accomplished by reacting the label carried by the binding partner with another substance, to form a detectable signal. When the label is an enzyme, as it frequently is, the substance is a substrate for the enzyme. The substrate for the enzyme either forms a visible color or changes color. Measuring the change or amount of color provides a measure of the produced complex, and hence of the analyte.
There is a need for quick, accurate, simple assays that can be performed by laboratory personnel as well as by non-technical personnel outside of a laboratory setting to test biological fluids of organisms to determine the presence of biological analytes such as immunoglobulins and immunocomplexes in the blood that are associated with or indicative of food, food additive or chemical allergies; or cancer and autoimmune disease.
All publications, scientific, patent or otherwise, referenced herein are hereby incorporated by reference in their entirety for all purposes.